Bone implants and methods comprising demineralized bone material

ABSTRACT

Bone implant compositions and methods are provide that have an outer surface, the outer surface comprising demineralized bone and having a cavity disposed in the outer surface, the cavity having a demineralized bone material coated therein; and an inner surface of the bone implant comprising cortical bone, the inner surface contacting the outer surface. The bone implant compositions and methods provided are osteoinductive and allow rapid bone fusion.

This application is a divisional application of U.S. patent applicationSer. No. 13/465,122 filed on May 7, 2012, entitled “BONE IMPLANTS ANDMETHODS COMPRISING DEMINERALIZED BONE MATERIAL”. This entire disclosureis incorporated herein by reference into the present disclosure.

BACKGROUND

The rapid and effective repair of bone defects caused by injury,disease, wounds, or surgery is a goal of orthopedic surgery. Toward thisend, a number of bone implants have been used or proposed for use in therepair of bone defects. The biological, physical, and mechanicalproperties of the bone implants are among the major factors influencingtheir suitability and performance in various orthopedic applications.

Bone implants are used to repair bone that has been damaged by disease,trauma, or surgery. Bone implants may be utilized when healing isimpaired in the presence of certain drugs or in disease states such asdiabetes, when a large amount of bone or disc material is removed duringsurgery, or when bone fusion is needed to create stability. In sometypes of spinal fusion, for example, bone implants are used to replacethe cushioning disc material between the vertebrae.

One type of bone implant is the bone graft. Typically, bone graft (e.g.,osteograft) materials may include both synthetic and natural bone.Natural bone may be taken from the graft recipient (autograft) or may betaken from another source (allograft), such as a cadaver, or(xenograft), such as bovine. Autografts have advantages such asdecreased immunogenicity and greater osteoinductive potential, but therecan also be problems with donor site morbidity and limited supply ofsuitable bone for grafting. On the other hand, allografts are availablein greater supply and can be stored for years. However, allografts tendto be less osteoinductive.

Osteoconduction and osteoinduction both contribute to bone formation. Agraft material is osteoconductive if it provides a structural frameworkor microscopic and macroscopic scaffolding for cells and cellularmaterials that are involved in bone formation (e.g., osteoclasts,osteoblasts, vasculature, mesenchymal cells).

Osteoinductive material, on the other hand, stimulates differentiationof host mesenchymal cells into chondroblasts and osteoblasts. Naturalbone allograft materials can comprise either cortical or cancellousbone. A distinguishing feature of cancellous bone is its high level ofporosity relative to that of cortical bone, providing more free surfacesand more of the cellular constituents that are retained on thesesurfaces. It provides both an osteoinductive and osteoconductive graftmaterial, but generally does not have significant load-bearing capacity.Optimal enhancement of bone formation is generally thought to require aminimum threshold quantity of cancellous bone, however. Cortical(compact) bone has greater strength or load-bearing capacity thancancellous bone, but is less osteoconductive. In humans for example,only approximately twenty percent of large cortical allografts arecompletely incorporated at five years. Delayed or incompleteincorporation may allow micromotion, leading to host bone resorptionaround the allograft. A more optimal bone graft material would combinesignificant load-bearing capacity with both osteoinductive andosteoconductive properties, and much effort has been directed towarddeveloping such a graft material.

Some allografts comprise mammalian cadaver bone treated to remove allsoft tissue, including marrow and blood, and then textured to form amultiplicity of holes of selected size, spacing, and depth. The texturedbone section is then immersed and demineralized, for example, in adilute acid bath. Demineralizing the bone exposes osteoinductivefactors, but extensive demineralization of bone also decreases itsmechanical strength.

Allografts have also been formed of organic bone matrix withperforations that extend from one surface, through the matrix, to theother surface to provide continuous channels between opposite surfaces.The organic bone matrix is produced by partial or completedemineralization of natural bone. Although the perforations increase thescaffolding potential of the graft material and may be filled withosteoinductive material as well, perforating organic bone matrix throughthe entire diameter of the graft decreases its load-bearing capacity.

Partially-demineralized cortical bone constructs may besurface-demineralized to prepare the graft to be soaked in bonegrowth-promoting substances such as bone morphogenetic protein (BMP).Although this design allows greater exposure of the surrounding tissueto growth-promoting factors, the surface demineralization necessary toadhere a substantial amount of growth-promoting factors to the graftmaterial decreases the allograft's mechanical strength.

What is needed is a bone implant that combines the osteoinductive andosteoconductive properties of cancellous bone with the load-bearingcapacity provided by cortical allograft materials. Compositions andmethods are needed that facilitate bone remodeling and new bone growth,and integration of the bone implant (e.g., allograft) into host bone.

SUMMARY

Compositions and methods are provided that facilitate bone remodelingand new bone growth, and integration of the bone implant (e.g.,allograft) into host bone. In some embodiments, the bone implantprovides an improved surface between the dense cortical allograft and apatient's host bone to facilitate incorporation of the allograftconstruct and fusion to host bone. In some embodiments, the bone implantcomprises a cortical allograft that contains osteoinductivedemineralized bone matrix material on its surface to initiate the bonefusion process at the bone allograft surface interface and provide rapidbone bonding to the demineralized surface of the cortical allograft. Thebone implant of the current application, in some embodiments, is loadbearing and provides good mechanical strength.

In some embodiments, there is a bone implant comprising an outersurface, the outer surface having a cavity disposed in the outersurface, the cavity having a coating comprising demineralized bonematrix material disposed therein; and an inner surface of the boneimplant comprising cortical bone, the inner surface contacting the outersurface.

In some embodiments, a bone implant is provided comprising an outersurface, the outer surface comprising a demineralized bone matrixmaterial and having a cavity disposed in the outer surface, the cavityhaving a coating comprising demineralized bone matrix material disposedtherein; and an inner surface of the bone implant comprising corticalbone, the inner surface contacting the outer surface.

In some embodiments, there is an allograft comprising an outer surface,the outer surface comprising a demineralized bone matrix material andhaving a cavity disposed in the outer surface, the cavity having acoating comprising demineralized bone matrix material disposed therein;and an inner surface of the allograft comprising cortical bone, theinner surface contacting the outer surface and the cavity providing apassageway to the inner surface.

In some embodiments, there is a method for repairing a bone defect in apatient in need of such treatment, the method comprising inserting anallograft into the bone defect, the allograft comprising an outersurface, which comprises a demineralized bone matrix material and havinga cavity disposed in the outer surface, the cavity having a coatingcomprising demineralized bone matrix material disposed therein; and aninner surface of the allograft comprising cortical bone, the innersurface contacting the outer surface and the cavity providing apassageway to the inner surface.

Additional features and advantages of various embodiments will be setforth in part in the description that follows, and in part will beapparent from the description, or may be learned by practice of variousembodiments. The objectives and other advantages of various embodimentswill be realized and attained by means of the elements and combinationsparticularly pointed out in the description and appended claims.

BRIEF DESCRIPTION OF THE FIGURES

In part, other aspects, features, benefits and advantages of theembodiments will be apparent with regard to the following description,appended claims and accompanying drawings where:

FIG. 1 illustrates a side sectional view of an embodiment of a boneimplant that has been surface demineralized and has a plurality ofcavities disposed at discrete positions about its surface.

FIG. 2 illustrates a magnified side sectional view of an embodiment of abone implant that has been surface demineralized and has a cavitydisposed in its surface. The cavity is coated with demineralized bonematrix material shown as fibers.

FIG. 3 illustrates a cross-sectional view of the outer surface of anembodiment of a bone implant. In this view, the outside surface has notbeen surface demineralized and the implant has creases that allow thebone implant to be folded to place the implant at or in the bone defect.

FIG. 4 illustrates a cross-sectional view of the inner surface of anembodiment of a bone implant. In this view, a portion of the innersurface comprises demineralized bone. The inner surface also compriseschannels comprising demineralized bone. In this view, a portion of theinside surface has not been surface demineralized and the implant hascreases that allow the bone implant to be folded to place the implant ator in the bone defect.

FIG. 5 illustrates a cross-sectional view of the inner surface of anembodiment of a bone implant. In this view, the inner surface comprisesconcave channels comprising demineralized bone.

FIG. 6 illustrates a side view of an embodiment of a bone implant in theform of a structural allograft that has been surface demineralized.

It is to be understood that the figures are not drawn to scale. Further,the relation between objects in a figure may not be to scale, and may infact have a reverse relationship as to size. The figures are intended tobring understanding and clarity to the structure of each object shown,and thus, some features may be exaggerated in order to illustrate aspecific feature of a structure.

DETAILED DESCRIPTION

For the purposes of this specification and appended claims, unlessotherwise indicated, all numbers expressing quantities of ingredients,percentages or proportions of materials, reaction conditions, and othernumerical values used in the specification and claims, are to beunderstood as being modified in all instances by the term “about.”Accordingly, unless indicated to the contrary, the numerical parametersset forth in the following specification and attached claims areapproximations that may vary depending upon the desired propertiessought to be obtained by the present application. At the very least, andnot as an attempt to limit the application of the doctrine ofequivalents to the scope of the claims, each numerical parameter shouldat least be construed in light of the number of reported significantdigits and by applying ordinary rounding techniques.

Notwithstanding that the numerical ranges and parameters setting forththe broad scope of the invention are approximations, the numerical areas precise as possible. Any numerical value, however, inherentlycontains certain errors necessarily resulting from the standarddeviation found in their respective testing measurements. Moreover, allranges disclosed herein are to be understood to encompass any and allsubranges subsumed therein. For example, a range of “1 to 10” includesany and all subranges between (and including) the minimum value of 1 andthe maximum value of 10, that is, any and all subranges having a minimumvalue of equal to or greater than 1 and a maximum value of equal to orless than 10, e.g., 5.5 to 10.

Additionally, unless defined otherwise or apparent from context, alltechnical and scientific terms used herein have the same meanings ascommonly understood by one of ordinary skill in the art to which thisinvention belongs.

Unless explicitly stated or apparent from context, the following termsare phrases have the definitions provided below:

Definitions

It is noted that, as used in this specification and the appended claims,the singular forms “a,” “an,” and “the,” include plural referents unlessexpressly and unequivocally limited to one referent. Thus, for example,reference to “an allograft” includes one, two, three or more allografts.

The term “biodegradable” includes that all or parts of the carrierand/or implant will degrade over time by the action of enzymes, byhydrolytic action and/or by other similar mechanisms in the human body.In various embodiments, “biodegradable” includes that the carrier and/orimplant can break down or degrade within the body to non-toxiccomponents after or while a therapeutic agent has been or is beingreleased. By “bioerodible” it is meant that the carrier and/or implantwill erode or degrade over time due, at least in part, to contact withsubstances found in the surrounding tissue, fluids or by cellularaction. By “bioabsorbable” or “bioresorbable” it is meant that thecarrier and/or implant will be broken down and absorbed within the humanbody, for example, by a cell or tissue. “Biocompatible” means that theallograft will not cause substantial tissue irritation or necrosis atthe target tissue site.

The term “mammal” refers to organisms from the taxonomy class“mammalian,” including but not limited to humans, other primates such aschimpanzees, apes, orangutans and monkeys, rats, mice, cats, dogs, cows,horses, etc.

“A “therapeutically effective amount” or “effective amount” is such thatwhen administered, the drug (e.g., growth factor) results in alterationof the biological activity, such as, for example, promotion of bone,cartilage and/or other tissue (e.g., vascular tissue) growth, inhibitionof inflammation, reduction or alleviation of pain, improvement in thecondition through inhibition of an immunologic response, etc. The dosageadministered to a patient can be as single or multiple doses dependingupon a variety of factors, including the drug's administeredpharmacokinetic properties, the route of administration, patientconditions and characteristics (sex, age, body weight, health, size,etc.), extent of symptoms, concurrent treatments, frequency of treatmentand the effect desired. In some embodiments the implant is designed forimmediate release. In other embodiments the implant is designed forsustained release. In other embodiments, the implant comprises one ormore immediate release surfaces and one or more sustained releasesurfaces.

The phrase “immediate release” is used herein to refer to one or moretherapeutic agent(s) that is introduced into the body and that isallowed to dissolve in or become absorbed at the location to which it isadministered, with no intention of delaying or prolonging thedissolution or absorption of the drug.

The phrases “sustained release” and “sustain release” (also referred toas extended release or controlled release) are used herein to refer toone or more therapeutic agent(s) that is introduced into the body of ahuman or other mammal and continuously or continually releases a streamof one or more therapeutic agents over a predetermined time period andat a therapeutic level sufficient to achieve a desired therapeuticeffect throughout the predetermined time period.

The terms “treating” and “treatment” when used in connection with adisease or condition refer to executing a protocol that may include abone repair procedure, where the bone implant and/or one or more drugsare administered to a patient (human, other normal or otherwise or othermammal), in an effort to alleviate signs or symptoms of the disease orcondition or immunological response. Alleviation can occur prior tosigns or symptoms of the disease or condition appearing, as well asafter their appearance. Thus, treating or treatment includes preventingor prevention of disease or undesirable condition. In addition,treating, treatment, preventing or prevention do not require completealleviation of signs or symptoms, does not require a cure, andspecifically includes protocols that have only a marginal effect on thepatient.

The term “bone,” as used herein, refers to bone that is cortical,cancellous or cortico-cancellous of autogenous, allogenic, xenogenic, ortransgenic origin.

The term “allograft” refers to a graft of tissue obtained from a donorof the same species as, but with a different genetic make-up from, therecipient, as a tissue transplant between two humans.

The term “autologous” refers to being derived or transferred from thesame individual's body, such as for example an autologous bone marrowtransplant.

The term “osteoconductive,” as used herein, refers to the ability of anon-osteoinductive substance to serve as a suitable template orsubstance along which bone may grow.

The term “osteoinductive,” as used herein, refers to the quality ofbeing able to recruit cells from the host that have the potential tostimulate new bone formation. Any material that can induce the formationof ectopic bone in the soft tissue of an animal is consideredosteoinductive.

The term “osteoinduction” refers to the ability to stimulate theproliferation and differentiation of pluripotent mesenchymal stem cells(MSCs). In endochondral bone formation, stem cells differentiate intochondroblasts and chondrocytes, laying down a cartilaginous ECM, whichsubsequently calcifies and is remodeled into lamellar bone. Inintramembranous bone formation, the stem cells differentiate directlyinto osteoblasts, which form bone through direct mechanisms.Osteoinduction can be stimulated by osteogenic growth factors, althoughsome ECM proteins can also drive progenitor cells toward the osteogenicphenotype.

The term “osteoconduction” refers to the ability to stimulate theattachment, migration, and distribution of vascular and osteogenic cellswithin the graft material. The physical characteristics that affect thegraft's osteoconductive activity include porosity, pore size, andthree-dimensional architecture. In addition, direct biochemicalinteractions between matrix proteins and cell surface receptors play amajor role in the host's response to the graft material.

The term “osteogenic” refers to the ability of a graft material toproduce bone independently. To have direct osteogenic activity, thegraft must contain cellular components that directly induce boneformation. For example, an allograft seeded with activated MSCs wouldhave the potential to induce bone formation directly, withoutrecruitment and activation of host MSC populations. Because manyosteoconductive allografts also have the ability to bind and deliverbioactive molecules, their osteoinductive potential will be greatlyenhanced.

The term “osteoimplant,” as used herein, refers to any bone-derivedimplant prepared in accordance with the embodiments of this disclosureand therefore is intended to include expressions such as bone membrane,bone graft, etc.

The term “patient” refers to a biological system to which a treatmentcan be administered. A biological system can include, for example, anindividual cell, a set of cells (e.g., a cell culture), an organ, or atissue. Additionally, the term “patient” can refer to animals,including, without limitation, humans.

The term “xenograft” refers to tissue or organs from an individual ofone species transplanted into or grafted onto an organism of anotherspecies, genus, or family.

The term “demineralized,” as used herein, refers to any materialgenerated by removing mineral material from tissue, e.g., bone tissue.In certain embodiments, the demineralized compositions described hereininclude preparations containing less than 5% calcium and preferably lessthan 1% calcium by weight. Partially demineralized bone (e.g.,preparations with greater than 5% calcium by weight but containing lessthan 100% of the original starting amount of calcium) is also consideredwithin the scope of the disclosure. In some embodiments, demineralizedbone has less than 95% of its original mineral content. Demineralized isintended to encompass such expressions as “substantially demineralized,”“partially demineralized,” and “fully demineralized.” In someembodiments, part or all of the surface of the bone can bedemineralized. For example, part or all of the surface of the allograftcan be demineralized to a depth of from about 100 to about 5000 microns,or about 150 microns to about 1000 microns. If desired, the outersurface of the intervertebral implant can be masked with an acidresistant coating or otherwise treated a to selectively demineralizeunmasked portions of the outer surface of the intervertebral implant sothat the surface demineralization is at discrete positions on theimplant.

The term “demineralized bone matrix,” as used herein, refers to anymaterial generated by removing mineral material from bone tissue. Insome embodiments, the DBM compositions as used herein includepreparations containing less than 5% calcium and preferably less than 1%calcium by weight. Partially demineralized bone (e.g., preparations withgreater than 5% calcium by weight but containing less than 100% of theoriginal starting amount of calcium) are also considered within thescope of the disclosure.

The term “superficially demineralized,” as used herein, refers tobone-derived elements possessing at least about 90 weight percent oftheir original inorganic mineral content, the expression “partiallydemineralized” as used herein refers to bone-derived elements possessingfrom about 8 to about 90 weight percent of their original inorganicmineral content and the expression “fully demineralized” as used hereinrefers to bone containing less than 8% of its original mineral context.

The terms “pulverized bone”, “powdered bone” or “bone powder” as usedherein, refers to bone particles of a wide range of average particlesize ranging from relatively fine powders to coarse grains and evenlarger chips.

The allograft can comprise bone fibers. Fibers include bone elementswhose average length to average thickness ratio or aspect ratio of thefiber is from about 50:1 to about 1000:1. In overall appearance thefibrous bone elements can be described as elongated bone fibers,threads, narrow strips, or thin sheets. Often, where thin sheets areproduced, their edges tend to curl up toward each other. The fibrousbone elements can be substantially linear in appearance or they can becoiled to resemble springs. In some embodiments, the elongated bonefibers are of irregular shapes including, for example, linear,serpentine or curved shapes. The elongated bone fibers are preferablydemineralized however some of the original mineral content may beretained when desirable for a particular embodiment.

Non-fibrous, as used herein, refers to elements that have an averagewidth substantially larger than the average thickness of the fibrousbone element or aspect ratio of less than from about 50:1 to about1000:1. In some embodiments, the non-fibrous bone elements are shaped ina substantially regular manner or specific configuration, for example,triangular prism, sphere, cube, cylinder and other regular shapes. Bycontrast, particles such as chips, shards, or powders possess irregularor random geometries. It should be understood that some variation indimension will occur in the production of the elements of thisapplication and elements demonstrating such variability in dimension arewithin the scope of this application and are intended to be understoodherein as being within the boundaries established by the expressions“mostly irregular” and “mostly regular”.

Reference will now be made in detail to certain embodiments of theinvention. While the invention will be described in conjunction with theillustrated embodiments, it will be understood that they are notintended to limit the invention to those embodiments. On the contrary,the invention is intended to cover all alternatives, modifications, andequivalents that may be included within the invention as defined by theappended claims.

The headings below are not meant to limit the disclosure in any way;embodiments under any one heading may be used in conjunction withembodiments under any other heading.

Demineralized Bone Allograft

Compositions and methods are provided that facilitate bone remodelingand new bone growth, and integration of the bone implant (e.g.,allograft) into host bone. In some embodiments, the bone implantprovides an improved surface between the dense cortical allograft and apatient's host bone to facilitate incorporation of the allograftconstruct and fusion to host bone. In some embodiments, the bone implantcomprises a cortical allograft that contains osteoinductivedemineralized bone matrix material on its surface to initiate the bonefusion process at the bone allograft surface interface and provide rapidbone bonding to the demineralized surface of the cortical allograft. Thebone implant of the current application, in some embodiments, is loadbearing and provides good mechanical strength.

In some embodiments, a bone implant is provided comprising an outersurface, the outer surface comprising a demineralized bone matrixmaterial and having a cavity disposed in the outer surface, the cavityhaving a coating comprising demineralized bone matrix material disposedtherein; and an inner surface of the bone implant comprising corticalbone, the inner surface contacting the outer surface.

In some embodiments, the bone implant comprises a structural allograftthat can be implanted between a dense cortical allograft and a patients'host bone to facilitate incorporation of the allograft construct andfusion to host bone. In this embodiment, this allograft can comprisecortical bone and can be surface demineralized and then coated with ademineralized powder. This interface will provide a therapeutic amountof osteoinductive DBM material to initiate the fusion process at theinterface and rapid bone bonding to the surface demineralized corticalallograft.

Current structural allograft implants can be made from dense corticalbone requiring significant time for the host bone to remodel theallograft interface surface via osteoclastic resorption and eventualdeposition of new bone into cutting cones into the allograft. Byemploying the bone implant of the current application that includessurface demineralized bone material, the fusion process can beaccelerated.

In some embodiments, the composite implant is configured to increase thesurface area contact of the allograft with the host bone, which willresult in faster fusion and incorporation of the composite implant intohost bone and ultimately a stronger fusion mass. In some embodiments,the allograft bone used in the implant is surface demineralization toincrease its osteoinductivity and fusion with the host bone. In someembodiments, the implant is designed so that the majority of themechanical load is carried by the allograft that comprises corticalbone.

In some embodiments, the portion of the allograft that is notdemineralized comprises load bearing and/or higher compressive strengthallograft material. In some embodiments, the portion of the allograftthat is not load bearing comprises demineralized bone material that alsohas a low compressive strength.

In some embodiments, the implant device contacts host bone and theimplant device comprises from about 1% to about 30% or from about 5% toabout 25% by weight of demineralized bone material.

In some embodiments, the bone allograft material comprises demineralizedbone matrix fibers and demineralized bone matrix powder in a ratio of25:75 to about 75:25 fibers to chips

In some embodiments, the surface demineralization provides a moreconducive surface for the demineralized bone to attach to via bothfriction and cohesive binding of collagen/protein compositions to theallograft. The healing process also exposes some of the inherent bonegrowth factors in the cortical allograft material to further facilitateremodeling and new bone formation. The surface demineralization of theallograft and/or DBM powder and/or fiber provides an easier route ofentry for bone remodeling to occur in the cortical allograft bonefurther facilitating faster fusion.

Demineralized bone matrix (DBM) is demineralized allograft bone withosteoinductive activity. DBM is prepared by acid extraction of allograftbone, resulting in loss of most of the mineralized component butretention of collagen and noncollagenous proteins, including growthfactors. DBM does not contain osteoprogenitor cells, but the efficacy ofa demineralized bone matrix as a bone-graft substitute or extender maybe influenced by a number of factors, including the sterilizationprocess, the carrier, the total amount of bone morphogenetic protein(BMP) present, and the ratios of the different BMPs present. DBMincludes demineralized pieces of cortical bone to expose theosteoinductive proteins contained in the matrix. These activateddemineralized bone particles are usually added to a substrate or carrier(e.g. glycerol or a polymer). DBM is mostly an osteoinductive product,but lacks enough induction to be used on its own in challenging healingenvironments such as posterolateral spine fusion.

Various designs are contemplated. The surface demineralization can bestrategically located and depth varied to optimize the incorporationprocess. For example it could be limited to certain areas on theinterface so as not to lead to allograft subsidence. Alternatively itcould be limited to the inner and outer surfaces to preferentiallyfacilitate allograft incorporation to those surfaces first. For example,part or all of the surface of the allograft can be demineralized to adepth of from about 50 to about 5000 microns, or about 100 microns toabout 1000 microns or about 2000 microns. If desired, the outer surfaceof the intervertebral implant can be masked with an acid resistantcoating or otherwise treated to selectively demineralize unmaskedportions of the outer surface of the intervertebral implant so that thesurface demineralization is at discrete positions on the implant.

Regarding coating with demineralized bone, it too can be strategicallylocated and depth varied to optimize the incorporation process. Forexample it could be limited to friction groves in the cortical allograftsurface so a thicker coating can be applied to accelerate new boneformation. One embodiment includes the use of concave groves filled withDBM since osteoblast preferentially attached to concave surfaces andthis is where bone formation typically occurs first. Alternatively itcould be limited to the inner and outer surfaces to preferentiallyfacilitate allograft incorporation to those surfaces first. The DBM isapplied to the surface demineralized surface and allowed to bind via airdrying, or alternatively freeze dried, heat drying, or a mild chemicalcrosslinking agent or adhesive can be used. The form of the DBM is canbe chips, shards, powders, fibers or a combination thereof, which areosteoinductive. These can be filled into the cavities of the allograftor one or more of them can be disposed on the allograft surface.Therefore, in some embodiments, the allograft can have both surfacedemineralization and DBM chips, shards, powders, or a combinationthereof disposed on its surface.

In one embodiment, DBM powder can range in average particle size fromabout 0.0001 to about 1.2 cm and from about 0.002 to about 1 cm. Thebone powder can be obtained from cortical, cancellous and/orcorticocancellous allogenic or xenogenic bone tissue. In general,allogenic bone tissue is preferred as the source of the bone powder.

In some embodiments, the coating thickness of DBM powder and/or fibersmay be thin, for example, from about 5, 10, 15, 20, 25, 30, 35, 40, 45or 50 microns to thicker coatings 60, 65, 70, 75, 80, 85, 90, 95, 100microns. In some embodiments, the range of the coating ranges from about5 microns to about 250 microns or 5 microns to about 200 microns.

According to some embodiments of the disclosure, the demineralized bonematrix may comprise demineralized bone matrix fibers and/ordemineralized bone matrix chips. In some embodiments, the demineralizedbone matrix may comprise demineralized bone matrix fibers anddemineralized bone matrix chips in a 30:60 ratio. According to oneembodiment of the disclosure, the bone composite comprises a bonepowder, a polymer and a demineralized bone. In different embodiments ofthe disclosure, bone powder content can range from about 5% to about 90%w/w, polymer content can range from about 5% to about 90% w/w, anddemineralized bone particles content comprises the reminder of thecomposition. Preferably, the demineralized bone particles comprise fromabout 20% to about 40% w/w while the polymer and the bone powdercomprise each from about 20% to about 60% w/w of the composition. Thebone graft materials of the present disclosure include those structuresthat have been modified in such a way that the original chemical forcesnaturally present have been altered to attract and bind molecules,including, without limitation, growth factors and/or cells, includingcultured cells.

Namely, the demineralized allograft bone material may be furthermodified such that the original chemical forces naturally present havebeen altered to attract and bind growth factors, other proteins andcells affecting osteogenesis, osteoconduction and osteoinduction. Forexample, a demineralized allograft bone material may be modified toprovide an ionic gradient to produce a modified demineralized allograftbone material, such that implanting the modified demineralized allograftbone material results in enhanced ingrowth of host bone.

In one embodiment an ionic force change agent may be applied to modifythe demineralized allograft bone material. The demineralized allograftbone material may comprise, e.g., a demineralized bone matrix (DBM)comprising fibers, particles and any combination of thereof. Accordingto another embodiment, a bone graft structure may be used whichcomprises a composite bone, which includes a bone powder, a polymer anda demineralized bone.

The ionic force change agent may be applied to the entire demineralizedallograft bone material or to selected portions/surfaces thereof.

The ionic force change agent may be a binding agent, which modifies thedemineralized allograft bone material or bone graft structure to bindmolecules, such as, for example, DBM, growth factors, or cells, such as,for example, cultured cells, or a combination of molecules and cells. Inthe practice of the disclosure the growth factors include but are notlimited to BMP-2, rhBMP-2, BMP-4, rhBMP-4, BMP-6, rhBMP-6, BMP-7 (OP-1),rhBMP-7, GDF-5, LIM mineralization protein, platelet derived growthfactor (PDGF), transforming growth factor-β (TGF-β), insulin-relatedgrowth factor-I (IGF-I), insulin-related growth factor-II (IGF-II),fibroblast growth factor (FGF), beta-2-microglobulin (BDGF II), andrhGDF-5. A person of ordinary skill in the art will appreciate that thedisclosure is not limited to growth factors only. Other molecules canalso be employed in the disclosure. For example, tartrate-resistant acidphosphatase, which is not a growth factor, may also be used in thedisclosure.

An adhesive may be applied to the DBM powders and/or fibers. Theadhesive material may comprise polymers having hydroxyl, carboxyl,and/or amine groups. In some embodiments, polymers having hydroxylgroups include synthetic polysaccharides, such as for example, cellulosederivatives, such as cellulose ethers (e.g., hydroxypropylcellulose). Insome embodiments, the synthetic polymers having a carboxyl group, maycomprise poly(acrylic acid), poly(methacrylic acid), poly(vinylpyrrolidone acrylic acid-N-hydroxysuccinimide), and poly(vinylpyrrolidone-acrylic acid-acrylic acid-N-hydroxysuccinimide) terpolymer.For example, poly(acrylic acid) with a molecular weight greater than250,000 or 500,000 may exhibit particularly good adhesive performance.In some embodiments, the adhesive can be a polymer having a molecularweight of about 2,000 to about 5,000, or about 10,000 to about 20,000 orabout 30,000 to about 40,000.

In some embodiments, the adhesive can comprise imido ester,p-nitrophenyl carbonate, N-hydroxysuccinimide ester, epoxide,isocyanate, acrylate, vinyl sulfone, orthopyridyl-disulfide, maleimide,aldehyde, iodoacetamide or a combination thereof. In some embodiments,the adhesive material can comprise at least one of fibrin, acyanoacrylate (e.g., N-butyl-2-cyanoacrylate, 2-octyl-cyanoacrylate,etc.), a collagen-based component, a glutaraldehyde glue, a hydrogel,gelatin, an albumin solder, and/or a chitosan adhesives. In someembodiments, the hydrogel comprises acetoacetate esters crosslinked withamino groups or polyethers as mentioned in U.S. Pat. No. 4,708,821. Insome embodiments, the adhesive material can comprise poly(hydroxylic)compounds derivatized with acetoacetate groups and/or polyaminocompounds derivatized with acetoacetamide groups by themselves or thecombination of these compounds crosslinked with an amino-functionalcrosslinking compounds.

The adhesive can be a solvent based adhesive, a polymer dispersionadhesive, a contact adhesive, a pressure sensitive adhesive, a reactiveadhesive, such as for example multi-part adhesives, one part adhesives,heat curing adhesives, moisture curing adhesives, or a combinationthereof or the like. The adhesive can be natural or synthetic or acombination thereof.

Contact adhesives are used in strong bonds with high shear-resistance.Pressure sensitive adhesives form a bond by the application of lightpressure to bind the adhesive with the target tissue site, cannulaand/or expandable member. In some embodiments, to have the device adhereto the target tissue site, pressure is applied in a directionsubstantially perpendicular to a surgical incision.

Multi-component adhesives harden by mixing two or more components, whichchemically react. This reaction causes polymers to cross-link intoacrylics, urethanes, and/or epoxies. There are several commercialcombinations of multi-component adhesives in use in industry. Some ofthese combinations are: polyester resin-polyurethane resin;polyols-polyurethane resin, acrylic polymers-polyurethane resins or thelike. The multi-component resins can be either solvent-based orsolvent-less. In some embodiments, the solvents present in the adhesivesare a medium for the polyester or the polyurethane resin. Then thesolvent is dried during the curing process.

In some embodiments, the adhesive can be a one-part adhesive. One-partadhesives harden via a chemical reaction with an external energy source,such as radiation, heat, and moisture. Ultraviolet (UV) light curingadhesives, also known as light curing materials (LCM), have becomepopular within the manufacturing sector due to their rapid curing timeand strong bond strength. Light curing adhesives are generally acrylicbased. The adhesive can be a heat-curing adhesive, where when heat isapplied (e.g., body heat), the components react and cross-link. Thistype of adhesive includes epoxies, urethanes, and/or polyimides. Theadhesive can be a moisture curing adhesive that cures when it reactswith moisture present (e.g., bodily fluid) on the substrate surface orin the air. This type of adhesive includes cyanoacrylates or urethanes.The adhesive can have natural components, such as for example, vegetablematter, starch (dextrin), natural resins or from animals e.g. casein oranimal glue. The adhesive can have synthetic components based onelastomers, thermoplastics, emulsions, and/or thermosets includingepoxy, polyurethane, cyanoacrylate, or acrylic polymers.

The allograft provides a matrix for the cells to guide the process oftissue formation in vivo in three dimensions. The morphology of theallograft guides cell migration and cells are able to migrate into orover the allograft, respectively. The cells then are able to proliferateand synthesize new tissue and form bone and/or cartilage. In someembodiments, one or more allografts are stacked on one or morebiodegradable carriers.

In some embodiments, the allograft comprises a plurality of pores. Insome embodiments, at least 10% of the pores are between about 10micrometers and about 500 micrometers at their widest points. In someembodiments, at least 20% of the pores are between about 50 micrometersand about 150 micrometers at their widest points. In some embodiments,at least 30% of the pores are between about 30 micrometers and about 70micrometers at their widest points. In some embodiments, at least 50% ofthe pores are between about 10 micrometers and about 500 micrometers attheir widest points. In some embodiments, at least 90% of the pores arebetween about 50 micrometers and about 150 micrometers at their widestpoints. In some embodiments, at least 95% of the pores are between about100 micrometers and about 250 micrometers at their widest points. Insome embodiments, 100% of the pores are between about 10 micrometers andabout 300 micrometers at their widest points.

In some embodiments, the allograft has a porosity of at least about 30%,at least about 50%, at least about 60%, at least about 70%, at leastabout 90%. The pore may support ingrowth of cells, formation orremodeling of bone, cartilage and/or vascular tissue.

The allograft in addition to bone and/or demineralized bone may comprisenatural and/or synthetic material. For example, the allograft maycomprise poly (alpha-hydroxy acids), poly (lactide-co-glycolide) (PLGA),polylactide (PLA), polyglycolide (PG), polyethylene glycol (PEG)conjugates of poly (alpha-hydroxy acids), polyorthoesters (POE),polyaspirins, polyphosphagenes, collagen, hydrolyzed collagen, gelatin,hydrolyzed gelatin, fractions of hydrolyzed gelatin, elastin, starch,pre-gelatinized starch, hyaluronic acid, chitosan, alginate, albumin,fibrin, vitamin E analogs, such as alpha tocopheryl acetate, d-alphatocopheryl succinate, D,L-lactide, or L-lactide, -caprolactone,dextrans, vinylpyrrolidone, polyvinyl alcohol (PVA), PVA-g-PLGA,PEGT-PBT copolymer (polyactive), methacrylates, poly(N-isopropylacrylamide), PEO-PPO-PEO (pluronics), PEO-PPO-PAAcopolymers, PLGA-PEO-PLGA, PEG-PLG, PLA-PLGA, poloxamer 407,PEG-PLGA-PEG triblock copolymers, SAIB (sucrose acetate is obutyrate),polydioxanone, methylmethacrylate (MMA), MMA and N-vinylpyrrolidone,polyamide, oxycellulose, copolymer of glycolic acid and trimethylenecarbonate, polyesteramides, polyetheretherketone,polymethylmethacrylate, or combinations thereof.

In some embodiments, the allograft may comprise a resorbable ceramic(e.g., hydroxyapatite, tricalcium phosphate, bioglasses, calciumsulfate, etc.) tyrosine-derived polycarbonate poly(DTE-co-DT carbonate),in which the pendant group via the tyrosine—an amino acid—is either anethyl ester (DTE) or free carboxylate (DT) or combinations thereof.

In some embodiments, the allograft may comprises collagen. Exemplarycollagens include human or non-human (bovine, ovine, and/or porcine), aswell as recombinant collagen or combinations thereof. Examples ofsuitable collagen include, but are not limited to, human collagen typeI, human collagen type II, human collagen type III, human collagen typeIV, human collagen type V, human collagen type VI, human collagen typeVII, human collagen type VIII, human collagen type IX, human collagentype X, human collagen type XI, human collagen type XII, human collagentype XIII, human collagen type XIV, human collagen type XV, humancollagen type XVI, human collagen type XVII, human collagen type XVIII,human collagen type XIX, human collagen type XXI, human collagen typeXXII, human collagen type XXIII, human collagen type XXIV, humancollagen type XXV, human collagen type XXVI, human collagen type XXVII,and human collagen type XXVIII, or combinations thereof. Collagenfurther may comprise hetero- and homo-trimers of any of theabove-recited collagen types. In some embodiments, the collagencomprises hetero- or homo-trimers of human collagen type I, humancollagen type II, human collagen type III, or combinations thereof.

In some embodiments, the allograft may be seeded with harvested bonecells and/or bone tissue, such as for example, cortical bone, autogenousbone, allogenic bones and/or xenogenic bone. In some embodiments, theallograft may be seeded with harvested cartilage cells and/or cartilagetissue (e.g., autogenous, allogenic, and/or xenogenic cartilage tissue).For example, before insertion into the target tissue site, the allograftcan be wetted with the graft bone tissue/cells, usually with bonetissue/cells aspirated from the patient, at a ratio of about 3:1, 2:1,1:1, 1:3 or 1:2 by volume. The bone tissue/cells are permitted to soakinto the allograft provided, and the allograft may be kneaded by hand,thereby obtaining a pliable consistency that may subsequently be packedinto the osteochondral defect.

The allograft may contain an inorganic material, such as an inorganicceramic and/or bone substitute material. Exemplary inorganic materialsor bone substitute materials include but are not limited to aragonite,dahlite, calcite, amorphous calcium carbonate, vaterite, weddellite,whewellite, struvite, urate, fenihydrate, francolite, monohydrocalcite,magnetite, goethite, dentin, calcium carbonate, calcium sulfate, calciumphosphosilicate, sodium phosphate, calcium aluminate, calcium phosphate,hydroxyapatite, alpha-tricalcium phosphate, dicalcium phosphate,β-tricalcium phosphate, tetracalcium phosphate, amorphous calciumphosphate, octacalcium phosphate, BIOGLASS™, fluoroapatite,chlorapatite, magnesium-substituted tricalcium phosphate, carbonatehydroxyapatite, substituted forms of hydroxyapatite (e.g.,hydroxyapatite derived from bone may be substituted with other ions suchas fluoride, chloride, magnesium sodium, potassium, etc.), orcombinations or derivatives thereof.

In some embodiments, the allograft has a density of between about 1.6g/cm³, and about 0.05 g/cm³. In some embodiments, the allograft has adensity of between about 1.1 g/cm³, and about 0.07 g/cm³. For example,the density may be less than about 1 g/cm³, less than about 0.7 g/cm³,less than about 0.6 g/cm³, less than about 0.5 g/cm³, less than about0.4 g/cm³, less than about 0.3 g/cm³, less than about 0.2 g/cm³, or lessthan about 0.1 g/cm³.

The shape of the allograft may be tailored to the site at which it is tobe situated. For example, it may be in the shape of a morsel, a plug, apin, a peg, a cylinder, a block, a wedge, ring, a sheet, etc.

In some embodiments, the allograft may be made by injection molding,compression molding, blow molding, thermoforming, die pressing, slipcasting, electrochemical machining, laser cutting, water-jet machining,electrophoretic deposition, powder injection molding, sand casting,shell mold casting, lost tissue scaffold casting, plaster-mold casting,ceramic-mold casting, investment casting, vacuum casting, permanent-moldcasting, slush casting, pressure casting, die casting, centrifugalcasting, squeeze casting, rolling, forging, swaging, extrusion,shearing, spinning, powder metallurgy compaction or combinationsthereof.

In some embodiments, a therapeutic agent may be disposed on or in theallograft by hand, electrospraying, ionization spraying or impregnating,vibratory dispersion (including sonication), nozzle spraying,compressed-air-assisted spraying, brushing and/or pouring. For example,a growth factor such as rhBMP-2 may be disposed on or in the allograft.

In some embodiments, the allograft may comprise sterile and/orpreservative free material.

In applying the allograft to the bone defect, the bone defect site inneed of treatment will be surgically prepared for receipt of theallograft. This preparation can include excision of bone at the site tocreate a hole or void in which the allograft will be received. Tissueremoval can be conducted in any suitable manner including for instancedrilling and/or punching, typically in a direction substantiallyperpendicular to the bone defect site, to create a recipient hole orvoid having a depth approximating that of the allograft to be implanted.

In some embodiments, the depth of the allograft can be for example, 1mm, 2 mm, 3 mm, 4 mm, 5 mm, 6 mm, 7 mm, 8 mm, 9 mm, 10 mm, 11 mm, 12 mm,13 mm, 14 mm, 15 mm, 16 mm, 17 mm, 18 mm, 19 mm, or 20 mm.

In some embodiments, the opening for receiving the allograft will becreated using a drill or punch having a circular cross-section. In someembodiments, multiple, overlapping passes with the drill or punch can bemade, in order to create an opening having a cross-section defined bymultiple, intersecting circular arcs. A chisel can then be used to shapethe recipient hole to receive the osteochondral implant.

In some embodiments, the allograft can include DBM particles, and/orcells (e.g., bone, chondrogenic cells and/or tissue) seeded or attachedto it.

In some embodiments, a small amount of biologic glue can be applied intothe hole or used to attached the DBM particles to the allograft, whichis also surface demineralized. Suitable organic glues include TISSEEL®or TISSUCOL® (fibrin based adhesive; Immuno AG, Austria), AdhesiveProtein (Sigma Chemical, USA), Dow Corning Medical Adhesive B (DowCorning, USA), fibrinogen thrombin, elastin, collagen, alginate,demineralized bone matrix, casein, albumin, keratin or the like. Acomposite fibrin glue can be mixed with milled cartilage from forexample, a bovine fibrinogen (e.g., SIGMA F-8630), thrombin (e.g., SIGMAT-4648) and aprotinin (e.g., SIGMA A6012. Also, human derivedfibrinogen, thrombin and aprotinin can be used.

In some embodiments, a method for repairing a bone defect is in apatient in need of such treatment is provided, the method comprisinginserting an allograft into the bone defect, the allograft comprising anouter surface, which comprises a demineralized bone matrix material andhaving a cavity disposed in the outer surface, the cavity having acoating comprising demineralized bone matrix material disposed therein;and an inner surface of the allograft comprising cortical bone, theinner surface contacting the outer surface and the cavity providing apassageway to the inner surface.

In some embodiments, a method for repairing a bone defect is providedwhere the method comprises forming a hole at or near the bone defect andinserting the allograft into the bone defect, wherein the cavity iscompletely filled with demineralized bone powder or demineralized bonefibers.

Now referring to the figures, FIG. 1 illustrates a side sectional viewof an embodiment of a bone implant device, cortical allograft 10 isshown that has a first surface 30 and a second surface 20. Surfaces 20,30 comprise allograft bone 60. Allograft 10 includes cavities 40disposed in a uniform or discrete arrangement on the outer edges ofsurfaces 20, 30. The term ‘cavity’ includes and encompasses voids,apertures, bores, depressions, holes, indentations, grooves, channels,notches or the like. In some embodiments, cavities 40 may be provided inany of a variety of shapes in addition to the generally rectangularshape shown, including but not limited to generally circular, oblong,curved, triangular and other polygonal or non-polygonal shapes. Forexample, each cavity can comprise a shape that is triangular, pyramidal,square, rectangular, pentagonal, hexagonal, heptagonal, octagonal,U-shaped, V-shaped, W-shaped, concave, crescent, or a combinationthereof. Cavities 40 contact host bone with a demineralized bone portioncontaining demineralized bone powder, fibers, shards, chips, or the like50. The surface of the cavity 40 can, in some embodiments, be surfacedemineralized. In some embodiments, the cavities 40 can be configured tomate with a corresponding mating surface in host bone. Thiscorresponding mating surface in host bone, in some embodiments, can bemade in host bone, by for example, drilling, etching, chisel, or otherdevice. In this way the cavities can mate with the correspondingcavities 40 from the allograft. The cavity can comprise channels that asshown in FIG. 1 extend longitudinally in the outer surface and thesechannels are liner and parallel to each other. The channels allow, amongother things, cells to enter the allograft and contact the inner surfaceof the allograft. In some embodiments, the channels may be disposeduniformly or randomly throughout the demineralized portion of theallograft.

In some embodiments, the depth of the demineralized region of theallograft of FIG. 1 varies along the outer surface. In some embodiments,the outer surface of the allograft is selectively demineralized so as toavoid subsidence when the bone implant is implanted into a host bone ofa patient.

FIG. 2 illustrates a magnified side sectional view of an embodiment ofthe implant that has been surface demineralized 80 and has cavity 70disposed in its surface. Cavity 70 contains demineralized bone powder100. The demineralized bone powder 100 can be coated in or on the cavityusing a suitable adhesive, glue, binder, carrier, or in someembodiments, the demineralized bone powder can be agglomerated andpacked into cavity 70.

In some embodiments, the demineralized bone powder, fiber, shards,and/or chips can be attached to the cavity by drying, freeze drying,heat drying, or using a chemical crosslinking agent.

Cortical bone 90 is disposed on the surfaces of allograft 10,surrounding cavity 70. Therefore, in this embodiment, the allograft hasa surface demineralized portion 80 and demineralized bone powder 100disposed on the surface demineralized portion in cavity 70. The corticalbone will be load bearing and have compressive strength, while thedemineralized portion will be osteoinductive and less load bearing andhave less compressive strength than the cortical portion of theallograft.

In some embodiments, the cavities are predrilled and are partially orcompletely filled with plugs or inserts that are demineralized, orsurface demineralized. In some embodiments, the plugs or insertscomprise cortical bone that is surface demineralized and then the plugor insert is friction fit or placed into the cavity of the implant, theplug or insert stays in place by friction.

In some embodiments, the cavities can be drilled into the implant orsurrounding bone and then the demineralized plug or insert implantedinto the prepared cavity. In some embodiments, the plugs or inserts stayin the cavities easier if the plugs or insert are cored from freezedried demineralized bone and then press-fit rather than coringundemineralized or fully mineralized bone plugs or inserts,demineralizing them, press-fitting into the cavities and freeze dryingthe complete bone implant. Freeze drying causes the demineralized plugsto shrink and fall out of the cavities. In some embodiments, thedemineralized plug or insert is flush with the bone implant. In someembodiments, the demineralized plug or insert protrudes from the boneimplant. In some embodiments, the demineralized plug or insert is notflush with the bone implant, but is below its surface.

In some embodiments, the bone implant is designed to optimize contact ofthe demineralized bone with the host bone.

In some embodiments, the shape of the demineralized bone plug or insertmay be tailored to the site at which it is to be situated. For example,it may be in the shape of a morsel, a plug, a pin, a peg, a cylinder, ablock, a wedge, a sheet, etc. that can be implanted into the corticalbone implant, which itself can be surface demineralized.

FIG. 3 illustrates a cross-sectional view of the outer surface 110 of anembodiment of a bone implant. In this view, the outside surface 110 hasnot been surface demineralized. Instead, the inside surface includesdemineralized bone material 120. In this embodiment shown, the implantcomprises creases that allow the bone implant to be folded to place theimplant at or in the bone defect. All or portions of the allograft cancomprise a therapeutic agent disposed in or on the allograft. In someembodiments, the creases comprise a therapeutic agent that when foldedin, on or around the bone defect, the creases release the therapeuticagent.

FIG. 4 illustrates a cross-sectional view of the inner surface 130 of anembodiment of a bone implant. In this view, a portion of the innersurface 130 comprises demineralized bone, such as, for example, surfaces150, 160. The inner surface 130 also comprises channels 140 that containdemineralized bone powder, shards, fibers, chips, or combinationsthereof. In this view, a portion of the inside surface 130 has not beensurface demineralized and the implant has creases (also shown in FIG. 3)that allow the bone implant to be folded to place the implant at, on, orin the bone defect.

FIG. 5 illustrates a cross-sectional view of the inner surface of anembodiment of a bone implant. In this view, the inner surface 170comprises concave cavities 180 that comprise demineralized bone powder190. In some embodiments, the concave cavities 180 can be configured tomate with a corresponding mating surface in host bone. Thiscorresponding mating surface in host bone, in some embodiments, can bemade in host bone, by for example, drilling, etching, chisel, or otherdevice. In this way the cavities can mate with the correspondingcavities 180 from the allograft.

FIG. 6 illustrates a side view of an embodiment of a bone implant in theform of a structural allograft ring 200 that has been surfacedemineralized 210. The surface is coated with demineralized bone fibers220 and growth factors 230 can be added. The allograft ring 200 includesa non-demineralized core 240. This type of implant can be amendable toinsertion between intervertebral discs, and/or for repair of the facetjoint.

The bone implant device may also include mechanisms or features forreducing and/or preventing slippage or migration of the device duringinsertion. For example, one or more surfaces of the implant may includeprojections such as ridges or teeth (not shown) for increasing thefriction between the implant and the adjacent contacting surfaces of thebone so to prevent movement of the implant after introduction to adesired location.

According to some embodiments, fusion may be facilitated or augmented byintroducing or positioning various osteoinductive materials within thecavities in the implant device. Such osteoinductive materials may beintroduced before, during, or after insertion of the exemplary implantdevice, and may include (but are not necessarily limited to) autologousbone harvested from the patient receiving the implant device, boneallograft, bone xenograft, any number of non-bone implants (e.g.ceramic, metallic, polymer), bone morphogenic protein, and/orbio-resorbable compositions.

In some embodiments, the bone implant may comprise an allograft portionthat is configured to be joined to another allograft portion or anon-allograft portion comprising a polymer. In this way, the implant canbe joined before it is implanted at or near the target site. The implantcan have mating surfaces comprising recesses and/or projections andreciprocating recesses and/or projections (e.g., joints) that allow theimplant to be assembled before implantation. Assembly can also include,for example, use of an adhesive material to join parts of the implanttogether and provide strong interlocking fit.

Growth Factors

In some embodiments, a growth factor and/or therapeutic agent may bedisposed on or in the allograft by hand, electrospraying, ionizationspraying or impregnating, vibratory dispersion (including sonication),nozzle spraying, compressed-air-assisted spraying, brushing and/orpouring. For example, a growth factor such as rhBMP-2 may be disposed onor in the allograft by the surgeon before the allograft is administeredor it may be available from the manufacturer beforehand.

The allograft may comprise at least one growth factor. These growthfactors include osteoinductive agents (e.g., agents that cause new bonegrowth in an area where there was none) and/or osteoconductive agents(e.g., agents that cause in growth of cells into and/or through theallograft). Osteoinductive agents can be polypeptides or polynucleotidescompositions. Polynucleotide compositions of the osteoinductive agentsinclude, but are not limited to, isolated Bone Morphogenetic Protein(BMP), Vascular Endothelial Growth Factor (VEGF), Connective TissueGrowth Factor (CTGF), Osteoprotegerin, Growth Differentiation Factors(GDFs), Cartilage Derived Morphogenic Proteins (CDMPs), LimMineralization Proteins (LMPs), Platelet derived growth factor, (PDGF orrhPDGF), Insulin-like growth factor (IGF) or Transforming Growth Factorbeta (TGF-beta) polynucleotides. Polynucleotide compositions of theosteoinductive agents include, but are not limited to, gene therapyvectors harboring polynucleotides encoding the osteoinductivepolypeptide of interest. Gene therapy methods often utilize apolynucleotide, which codes for the osteoinductive polypeptideoperatively linked or associated to a promoter or any other geneticelements necessary for the expression of the osteoinductive polypeptideby the target tissue. Such gene therapy and delivery techniques areknown in the art, (See, for example, International Publication No.WO90/11092, the disclosure of which is herein incorporated by referencein its entirety). Suitable gene therapy vectors include, but are notlimited to, gene therapy vectors that do not integrate into the hostgenome. Alternatively, suitable gene therapy vectors include, but arenot limited to, gene therapy vectors that integrate into the hostgenome.

In some embodiments, the polynucleotide is delivered in plasmidformulations. Plasmid DNA or RNA formulations refer to polynucleotidesequences encoding osteoinductive polypeptides that are free from anydelivery vehicle that acts to assist, promote or facilitate entry intothe cell, including viral sequences, viral particles, liposomeformulations, lipofectin, precipitating agents or the like. Optionally,gene therapy compositions can be delivered in liposome formulations andlipofectin formulations, which can be prepared by methods well known tothose skilled in the art. General methods are described, for example, inU.S. Pat. Nos. 5,593,972, 5,589,466, and 5,580,859, the disclosures ofwhich are herein incorporated by reference in their entireties.

Gene therapy vectors further comprise suitable adenoviral vectorsincluding, but not limited to for example, those described in U.S. Pat.No. 5,652,224, which is herein incorporated by reference.

Polypeptide compositions of the isolated osteoinductive agents include,but are not limited to, isolated Bone Morphogenetic Protein (BMP),Vascular Endothelial Growth Factor (VEGF), Connective Tissue GrowthFactor (CTGF), Osteoprotegerin, Growth Differentiation Factors (GDFs),Cartilage Derived Morphogenic Proteins (CDMPs), Lim MineralizationProteins (LMPs), Platelet derived growth factor, (PDGF or rhPDGF),Insulin-like growth factor (IGF) or Transforming Growth Factor beta(TGF-beta707) polypeptides. Polypeptide compositions of theosteoinductive agents include, but are not limited to, full lengthproteins, fragments or variants thereof.

Variants of the isolated osteoinductive agents include, but are notlimited to, polypeptide variants that are designed to increase theduration of activity of the osteoinductive agent in vivo. Preferredembodiments of variant osteoinductive agents include, but are notlimited to, full length proteins or fragments thereof that areconjugated to polyethylene glycol (PEG) moieties to increase theirhalf-life in vivo (also known as pegylation). Methods of pegylatingpolypeptides are well known in the art (See, e.g., U.S. Pat. No.6,552,170 and European Pat. No. 0,401,384 as examples of methods ofgenerating pegylated polypeptides). In some embodiments, the isolatedosteoinductive agent(s) are provided as fusion proteins. In oneembodiment, the osteoinductive agent(s) are available as fusion proteinswith the Fc portion of human IgG. In another embodiment, theosteoinductive agent(s) are available as hetero- or homodimers ormultimers. Examples of some fusion proteins include, but are not limitedto, ligand fusions between mature osteoinductive polypeptides and the Fcportion of human Immunoglobulin G (IgG). Methods of making fusionproteins and constructs encoding the same are well known in the art.

Isolated osteoinductive agents that are included within carrier aretypically sterile. In a non-limiting method, sterility is readilyaccomplished for example by filtration through sterile filtrationmembranes (e.g., 0.2 micron membranes or filters). In one embodiment,the isolated osteoinductive agents include one or more members of thefamily of Bone Morphogenetic Proteins (“BMPs”). BMPs are a class ofproteins thought to have osteoinductive or growth-promoting activitieson endogenous bone tissue, or function as pro-collagen precursors. Knownmembers of the BMP family include, but are not limited to, BMP-1, BMP-2,BMP-3, BMP-4, BMP-5, BMP-6, BMP-7, BMP-8, BMP-9, BMP-10, BMP-11, BMP-12,BMP-13, BMP-15, BMP-16, BMP-17, BMP-18 as well as polynucleotides orpolypeptides thereof, as well as mature polypeptides or polynucleotidesencoding the same.

BMPs utilized as osteoinductive agents comprise one or more of BMP-1;BMP-2; BMP-3; BMP-4; BMP-5; BMP-6; BMP-7; BMP-8; BMP-9; BMP-10; BMP-11;BMP-12; BMP-13; BMP-15; BMP-16; BMP-17; or BMP-18; as well as anycombination of one or more of these BMPs, including full length BMPs orfragments thereof, or combinations thereof, either as polypeptides orpolynucleotides encoding the polypeptide fragments of all of the recitedBMPs. The isolated BMP osteoinductive agents may be administered aspolynucleotides, polypeptides, full length protein or combinationsthereof.

In another embodiment, isolated osteoinductive agents includeosteoclastogenesis inhibitors to inhibit bone resorption of the bonetissue surrounding the site of implantation by osteoclasts. Osteoclastand osteoclastogenesis inhibitors include, but are not limited to,osteoprotegerin polynucleotides or polypeptides, as well as matureosteoprotegerin proteins, polypeptides or polynucleotides encoding thesame. Osteoprotegerin is a member of the TNF-receptor superfamily and isan osteoblast-secreted decoy receptor that functions as a negativeregulator of bone resorption. This protein specifically binds to itsligand, osteoprotegerin ligand (TNFSF11/OPGL), both of which are keyextracellular regulators of osteoclast development.

Osteoclastogenesis inhibitors further include, but are not limited to,chemical compounds such as bisphosphonate, 5-lipoxygenase inhibitorssuch as those described in U.S. Pat. Nos. 5,534,524 and 6,455,541 (thecontents of which are herein incorporated by reference in theirentireties), heterocyclic compounds such as those described in U.S. Pat.No. 5,658,935 (herein incorporated by reference in its entirety),2,4-dioxoimidazolidine and imidazolidine derivative compounds such asthose described in U.S. Pat. Nos. 5,397,796 and 5,554,594 (the contentsof which are herein incorporated by reference in their entireties),sulfonamide derivatives such as those described in U.S. Pat. No.6,313,119 (herein incorporated by reference in its entirety), oracylguanidine compounds such as those described in U.S. Pat. No.6,492,356 (herein incorporated by reference in its entirety).

In another embodiment, isolated osteoinductive agents include one ormore members of the family of Connective Tissue Growth Factors(“CTGFs”). CTGFs are a class of proteins thought to havegrowth-promoting activities on connective tissues. Known members of theCTGF family include, but are not limited to, CTGF-1, CTGF-2, CTGF-4polynucleotides or polypeptides thereof, as well as mature proteins,polypeptides or polynucleotides encoding the same.

In another embodiment, isolated osteoinductive agents include one ormore members of the family of Vascular Endothelial Growth Factors(“VEGFs”). VEGFs are a class of proteins thought to havegrowth-promoting activities on vascular tissues. Known members of theVEGF family include, but are not limited to, VEGF-A, VEGF-B, VEGF-C,VEGF-D, VEGF-E or polynucleotides or polypeptides thereof, as well asmature VEGF-A, proteins, polypeptides or polynucleotides encoding thesame.

In another embodiment, isolated osteoinductive agents include one ormore members of the family of Transforming Growth Factor-beta genes(“TGFbetas”). TGF-betas are a class of proteins thought to havegrowth-promoting activities on a range of tissues, including connectivetissues. Known members of the TGF-beta family include, but are notlimited to, TGF-beta-1, TGF-beta-2, TGF-beta-3, polynucleotides orpolypeptides thereof, as well as mature protein, polypeptides orpolynucleotides encoding the same.

In another embodiment, isolated osteoinductive agents include one ormore Growth Differentiation Factors (“GDFs”). Known GDFs include, butare not limited to, GDF-1, GDF-2, GDF-3, GDF-7, GDF-10, GDF-11, andGDF-15. For example, GDFs useful as isolated osteoinductive agentsinclude, but are not limited to, the following GDFs: GDF-1polynucleotides or polypeptides corresponding to GenBank AccessionNumbers M62302, AAA58501, and AAB94786, as well as mature GDF-1polypeptides or polynucleotides encoding the same. GDF-2 polynucleotidesor polypeptides corresponding to GenBank Accession Numbers BC069643,BC074921, Q9UK05, AAH69643, or AAH74921, as well as mature GDF-2polypeptides or polynucleotides encoding the same. GDF-3 polynucleotidesor polypeptides corresponding to GenBank Accession Numbers AF263538,BC030959, AAF91389, AAQ89234, or Q9NR23, as well as mature GDF-3polypeptides or polynucleotides encoding the same. GDF-7 polynucleotidesor polypeptides corresponding to GenBank Accession Numbers AB158468,AF522369, AAP97720, or Q7Z4P5, as well as mature GDF-7 polypeptides orpolynucleotides encoding the same. GDF-10 polynucleotides orpolypeptides corresponding to GenBank Accession Numbers BC028237 orAAH28237, as well as mature GDF-10 polypeptides or polynucleotidesencoding the same.

GDF-11 polynucleotides or polypeptides corresponding to GenBankAccession Numbers AF100907, NP_005802 or 095390, as well as matureGDF-11 polypeptides or polynucleotides encoding the same. GDF-15polynucleotides or polypeptides corresponding to GenBank AccessionNumbers BC008962, BC000529, AAH00529, or NP004855, as well as matureGDF-15 polypeptides or polynucleotides encoding the same.

In another embodiment, isolated osteoinductive agents include CartilageDerived Morphogenic Protein (CDMP) and Lim Mineralization Protein (LMP)polynucleotides or polypeptides. Known CDMPs and LMPs include, but arenot limited to, CDMP-1, CDMP-2, LMP-1, LMP-2, or LMP-3. CDMPs and LMPsuseful as isolated osteoinductive agents include, but are not limitedto, the following CDMPs and LMPs: CDMP-1 polynucleotides andpolypeptides corresponding to GenBank Accession Numbers NM_000557,U13660, NP_000548 or P43026, as well as mature CDMP-1 polypeptides orpolynucleotides encoding the same. CDMP-2 polypeptides corresponding toGenBank Accession Numbers or P55106, as well as mature CDMP-2polypeptides. LMP-1 polynucleotides or polypeptides corresponding toGenBank Accession Numbers AF345904 or AAK30567, as well as mature LMP-1polypeptides or polynucleotides encoding the same. LMP-2 polynucleotidesor polypeptides corresponding to GenBank Accession Numbers AF345905 orAAK30568, as well as mature LMP-2 polypeptides or polynucleotidesencoding the same. LMP-3 polynucleotides or polypeptides correspondingto GenBank Accession Numbers AF345906 or AAK30569, as well as matureLMP-3 polypeptides or polynucleotides encoding the same.

In another embodiment, isolated osteoinductive agents include one ormore members of any one of the families of Bone Morphogenetic Proteins(BMPs), Connective Tissue Growth Factors (CTGFs), Vascular EndothelialGrowth Factors (VEGFs), Osteoprotegerin or any of the otherosteoclastogenesis inhibitors, Growth Differentiation Factors (GDFs),Cartilage Derived Morphogenic Proteins (CDMPs), Lim MineralizationProteins (LMPs), or Transforming Growth Factor-betas (TGF-betas), TP508(an angiogenic tissue repair peptide), as well as mixtures orcombinations thereof.

In another embodiment, the one or more isolated osteoinductive agentsuseful in the bioactive formulation are selected from the groupconsisting of BMP-1, BMP-2, BMP-3, BMP-4, BMP-5, BMP-6, BMP-7, BMP-8,BMP-9, BMP-10, BMP-11, BMP-12, BMP-13, BMP-15, BMP-16, BMP-17, BMP-18,or any combination thereof; CTGF-1, CTGF-2, CGTF-3, CTGF-4, or anycombination thereof; VEGF-A, VEGF-B, VEGF-C, VEGF-D, VEGF-E, or anycombination thereof; GDF-1, GDF-2, GDF-3, GDF-7, GDF-10, GDF-11, GDF-15,or any combination thereof; CDMP-1, CDMP-2, LMP-1, LMP-2, LMP-3, and orcombination thereof; Osteoprotegerin; TGF-beta-1, TGF-beta-2,TGF-beta-3, or any combination thereof; or any combination of one ormore members of these groups.

The concentrations of growth factor can be varied based on the desiredlength or degree of osteogenic effects desired. Similarly, one of skillin the art will understand that the duration of sustained release of thegrowth factor can be modified by the manipulation of the compositionscomprising the sustained release formulation, such as for example,modifying the percent of allograft found within a sustained releaseformulation, microencapsulation of the formulation within polymers,including polymers having varying degradation times and characteristics,and layering the formulation in varying thicknesses in one or moredegradable polymers. These sustained release formulations can thereforebe designed to provide customized time release of growth factors thatsimulate the natural healing process.

In some embodiments, a statin may be used as the growth factor. Statinsinclude, but is not limited to, atorvastatin, simvastatin, pravastatin,cerivastatin, mevastatin (see U.S. Pat. No. 3,883,140, the entiredisclosure is herein incorporated by reference), velostatin (also calledsynvinolin; see U.S. Pat. Nos. 4,448,784 and 4,450,171 these entiredisclosures are herein incorporated by reference), fluvastatin,lovastatin, rosuvastatin and fluindostatin (Sandoz XU-62-320),dalvastain (EP Appln. Publn. No. 738510 A2, the entire disclosure isherein incorporated by reference), eptastatin, pitavastatin, orpharmaceutically acceptable salts thereof or a combination thereof. Invarious embodiments, the statin may comprise mixtures of (+)R and (−)-Senantiomers of the statin. In various embodiments, the statin maycomprise a 1:1 racemic mixture of the statin.

The growth factor may contain inactive materials such as bufferingagents and pH adjusting agents such as potassium bicarbonate, potassiumcarbonate, potassium hydroxide, sodium acetate, sodium borate, sodiumbicarbonate, sodium carbonate, sodium hydroxide or sodium phosphate;degradation/release modifiers; drug release adjusting agents;emulsifiers; preservatives such as benzalkonium chloride, chlorobutanol,phenylmercuric acetate and phenylmercuric nitrate, sodium bisulfate,sodium bisulfite, sodium thiosulfate, thimerosal, methylparaben,polyvinyl alcohol and phenylethyl alcohol; solubility adjusting agents;stabilizers; and/or cohesion modifiers. In some embodiments, the growthfactor may comprise sterile and/or preservative free material.

These above inactive ingredients may have multi-functional purposesincluding the carrying, stabilizing and controlling the release of thegrowth factor and/or other therapeutic agent(s). The sustained releaseprocess, for example, may be by a solution-diffusion mechanism or it maybe governed by an erosion-sustained process.

The amount of growth factor, e.g., bone morphogenic protein may besufficient to cause bone and/or cartilage growth. In some embodiments,the growth factor is rhBMP-2 and is contained in one or more carriers inan amount of from 0.05 to 2 mg per cubic centimeter of the biodegradablecarrier. In some embodiments, the amount of rhBMP-2 morphogenic proteinis from 2.0 to 2.5 mg per cubic centimeter (cc) of the biodegradablecarrier.

In some embodiments, the growth factor is supplied in an aqueousbuffered solution. Exemplary aqueous buffered solutions include, but arenot limited to, TE, HEPES(2-[4-(2-hydroxyethyl)-1-piperazinyl]ethanesulfonic acid), MES(2-morpholinoethanesulfonic acid), sodium acetate buffer, sodium citratebuffer, sodium phosphate buffer, a Tris buffer (e.g., Tris-HCL),phosphate buffered saline (PBS), sodium phosphate, potassium phosphate,sodium chloride, potassium chloride, glycerol, calcium chloride or acombination thereof. In various embodiments, the buffer concentrationcan be from about 1 mM to 100 mM.

In some embodiments, the BMP-2 is provided in a vehicle (including abuffer) containing sucrose, glycine, L-glutamic acid, sodium chloride,and/or polysorbate 80.

Additional Therapeutic Agents

The growth factors of the present application may be disposed on or inthe allograft and/or carrier with other therapeutic agents. For example,the growth factor may be disposed on or in the carrier byelectrospraying, ionization spraying or impregnating, vibratorydispersion (including sonication), nozzle spraying,compressed-air-assisted spraying, brushing and/or pouring.

Exemplary therapeutic agents include but are not limited to IL-1inhibitors, such Kineret® (anakinra), which is a recombinant,non-glycosylated form of the human inerleukin-1 receptor antagonist(IL-1Ra), or AMG 108, which is a monoclonal antibody that blocks theaction of IL-1. Therapeutic agents also include excitatory amino acidssuch as glutamate and aspartate, antagonists or inhibitors of glutamatebinding to NMDA receptors, AMPA receptors, and/or kainate receptors.Interleukin-1 receptor antagonists, thalidomide (a TNF-α releaseinhibitor), thalidomide analogues (which reduce TNF-α production bymacrophages), quinapril (an inhibitor of angiotensin II, whichupregulates TNF-α), interferons such as IL-11 (which modulate TNF-αreceptor expression), and aurin-tricarboxylic acid (which inhibitsTNF-α), may also be useful as therapeutic agents for reducinginflammation. It is further contemplated that where desirable apegylated form of the above may be used. Examples of still othertherapeutic agents include NF kappa B inhibitors such as antioxidants,such as dithiocarbamate, and other compounds, such as, for example,sulfasalazine.

Examples of therapeutic agents suitable for use also include, but arenot limited to an anti-inflammatory agent, analgesic agent, orosteoinductive growth factor or a combination thereof. Anti-inflammatoryagents include, but are not limited to, apazone, celecoxib, diclofenac,diflunisal, enolic acids (piroxicam, meloxicam), etodolac, fenamates(mefenamic acid, meclofenamic acid), gold, ibuprofen, indomethacin,ketoprofen, ketorolac, nabumetone, naproxen, nimesulide, salicylates,sulfasalazine [2-hydroxy-5-[-4-[C2-pyridinylamino)sulfonyl]azo]benzoicacid, sulindac, tepoxalin, and tolmetin; as well as antioxidants, suchas dithiocarbamate, steroids, such as cortisol, cortisone,hydrocortisone, fludrocortisone, prednisone, prednisolone,methylprednisolone, triamcinolone, betamethasone, dexamethasone,beclomethasone, fluticasone or a combination thereof.

Suitable analgesic agents include, but are not limited to,acetaminophen, bupivicaine, fluocinolone, lidocaine, opioid analgesicssuch as buprenorphine, butorphanol, dextromoramide, dezocine,dextropropoxyphene, diamorphine, fentanyl, alfentanil, sufentanil,hydrocodone, hydromorphone, ketobemidone, levomethadyl, mepiridine,methadone, morphine, nalbuphine, opium, oxycodone, papaveretum,pentazocine, pethidine, phenoperidine, piritramide, dextropropoxyphene,remifentanil, tilidine, tramadol, codeine, dihydrocodeine, meptazinol,dezocine, eptazocine, flupirtine, amitriptyline, carbamazepine,gabapentin, pregabalin, or a combination thereof.

Kits

In various embodiments, a kit is provided that may include additionalparts along with the allograft to be used to implant the allograft. Thekit may include the allograft in a first compartment. The secondcompartment may include a biodegradable carrier and the growth factorand any other instruments needed for the implanting the osteochondralimplant. A third compartment may include gloves, drapes, wound dressingsand other procedural supplies for maintaining sterility during theimplanting process, as well as an instruction booklet. A fourthcompartment may include additional tools for implantation (e.g., drills,drill bits, bores, punches, etc.). Each tool may be separately packagedin a plastic pouch that is radiation sterilized. A fifth compartment maycomprise an agent for radiographic imaging or the agent may be disposedon the allograft and/or carrier to monitor placement and tissue growth.A cover of the kit may include illustrations of the implanting procedureand a clear plastic cover may be placed over the compartments tomaintain sterility.

It will be apparent to those skilled in the art that variousmodifications and variations can be made to various embodimentsdescribed herein without departing from the spirit or scope of theteachings herein. Thus, it is intended that various embodiments coverother modifications and variations of various embodiments within thescope of the present teachings.

What is claimed is:
 1. A method for repairing a bone defect in a patientin need of such treatment, the method comprising inserting an allograftinto the bone defect, the allograft comprising an outer surface, whichcomprises a demineralized bone matrix material and having a cavitydisposed in the outer surface, the cavity having a coating comprisingdemineralized bone matrix material disposed therein; and an innersurface of the allograft comprising cortical bone, the inner surfacecontacting the outer surface and the cavity providing a passageway tothe inner surface, wherein the cavity comprises channels that extendlongitudinally in the outer surface and that are linear and parallel toeach other.
 2. A method for repairing a bone defect of claim 1, themethod comprising forming a space at or near the bone defect andinserting the allograft into the bone defect, wherein the cavity iscompletely filled with demineralized bone powder, demineralized bonefibers, a demineralized plug, or a demineralized insert.
 3. A method forrepairing a bone defect of claim 2, wherein at least one bioactive agentis added to the space.
 4. A method for repairing a bone defect of claim3, wherein the at least one bioactive agent comprises bone morphogeneticprotein (BMP), growth differentiation factor (GDF), LIM MineralizationProtein (LMP), TP508 (an angiogenic tissue repair peptide), bone marrowaspirate, concentrated bone marrow aspirate, platelets, mesenchymalcells, antibiotics, anti-infective compositions, analgesic agents,anti-inflammatory agents, or combinations thereof.
 5. A bone implantaccording to claim 1, wherein the demineralized bone matrix materialcomprises from about 22% to about 30% by weight of the implant.
 6. Amethod for repairing a bone defect in a patient in need of suchtreatment, the method comprising inserting an allograft into the bonedefect, the allograft comprising an outer surface, which comprises ademineralized bone matrix material comprising from about 1% to about 30%by weight of the implant, the allograft having a cavity disposed in theouter surface, the cavity having a coating comprising demineralized bonematrix material disposed therein; and an inner surface of the allograftcomprising cortical bone, the inner surface contacting the outer surfaceand the cavity providing a passageway to the inner surface, wherein thedemineralized bone matrix material comprises demineralized bone matrixfibers and demineralized bone matrix chips in a ratio of 25:75 to 75:25,and the coating has a thickness from about 5 microns to about 250microns, and the cavity comprises channels that extend longitudinally inthe outer surface that are linear and parallel to each other.